Spinocerebellar Ataxia type 3 (SCA3), also known as Machado-Joseph disease, is a dominantly inherited, progressive ataxia for which there is no cure. It is one of at least nine neurodegenerative disorders caused by CAG repeats encoding polyglutamine expansions in the respective disease genes. The expansion in SCA3 confers a toxic property on the ATXN3 disease gene product, a nonessential protein known as ataxin-3. Accordingly, suppressing the expression of ATXN3 is a potentially powerful therapeutic strategy for SCA3. Using a YAC transgenic mouse model expressing the full length human ATXN3 disease gene, we propose to test the in vivo efficacy and safety of viral-mediated RNA interference (RNAi) as therapy for SCA3. Preliminary studies have optimized microRNA-like short hairpin RNAs (shRNAs) that effectively target human ATXN3. Aim 1 will test and compare the ability of recombinant adeno-associated viruses (AAV) expressing these shRNAs to suppress expression of mutant human ATXN3 in injected cerebellum of adult mice. Parallel studies in aim 1 will also assess the safety of RNAi reagents targeting ATXN3. In Aim 2, the RNAi AAVs tested and confirmed in aim 1 will be injected bilaterally in the cerebellum, the primary neuropathological target in SCA3. Long term efficacy of RNAi to suppress both ATXN3 expression and disease-related phenotypes will be assessed. Because our AAV reagents employ elements already used in human gene therapy trials, a successful anti- ATXN3 RNAi reagent that fulfills safety and efficacy measures will represent an immediate candidate for preclinical studies leading toward a human clinical trial. PUBLIC HEALTH RELEVANCE: The proposed research will test viral-mediated RNA interference as a therapeutic strategy for the neurodegenerative disease, spinocerebellar ataxia type 3, a currently untreatable, fatal disorder. Studies carried out in a mouse model of disease are anticipate to establish that RNAi may be an effective strategy in humans with the disease.